ABSTRACT Human leukocyte antigen (HLA) typing is an essential laboratory test to evaluate the donor-recipient compatibility before life-saving transplantations. In 2016 alone, HLA typing of donors and recipients guided over 30,000 organ and stem cell transplantations in the United States. Existing high-resolution HLA typing methods require a turnaround time of days to weeks. There has been an unmet clinical need for a rapid, high- resolution HLA typing assay to accelerate donor selection and organ allocation at increased precision. We propose to develop a kit with bundled software to enable rapid, high-resolution HLA typing by nanopore sequencing. The MinION device by Oxford Nanopore Technologies generates long sequence reads by passing DNA through nanopores while recording the current change in real time. It is an open platform that supports the development of rapid molecular assays, although the high error rate (~8%) has hindered its broad application in human genetic testing. The product to be developed will overcome this hurdle to enable MinION- based high-resolution HLA typing. The long-term goal of this project is to improve transplant outcomes and further our understanding of transplant immunology through this diagnostic innovation. Previously, our research team created the ATHLON bioinformatics pipeline and successfully typed the class I HLA genes at the key exon level using reads from the MinION device. Based on this work, we hypothesize that the ATHLON approach can be adapted to the typing of class II HLA genes by MinION sequencing, and the class I and II typing resolution can be increased by considering regions beyond key exons. In this Phase I study, we aim to demonstrate accurate class II HLA typing by MinION sequencing because a successful HLA typing product must be able to type both class I and II HLA genes. We also aim to demonstrate accurate class I and II HLA typing at a resolution beyond key exons by MinION sequencing within a graphical user interface. Upon meeting the Phase I milestones, the Phase II project will focus on streamlining the workflow to meet an 8-hour turnaround time, establishing quality metrics in the software, and performing a multicenter validation study to establish the robustness of the product. The proposed HLA typing product will shift the paradigms in the field of HLA diagnostics with a broad appeal to histocompatibility laboratories of different sizes. With a near-zero capital investment and a significant saving of hands-on time, we expect quick and widespread adoption of this technology in routine clinical HLA typing.